Cyclohex-1,4-ylenedimethanol

Endpoint summary

Description of key information

In a subchronic oral toxicity study, 1,4-cyclohexanedimethanol was administered via the drinking water to groups of male and female Crl:CD(SD)IGS BR rats (10 females/dose and 12 males/dose) at target concentrations of 0 (0 mg/mL), 0.4% (4.0 mg/mL), 0.8% (8.0 mg/mL), and 1.25% (12.5 mg/mL) for 13 weeks. One male rat each from the low- and high-dose groups were found dead or euthanized in extremis before study termination. Clinical abnormalities related to test substance exposure included minimal to severe hematuria and/or brown/red discoloration of the urine, primarily in high-dose group animals. Softened or reduced feces occurred in animals in all groups but severity and incidence were slightly greater among treated groups. Other abnormal signs were observed occasionally in a few treated animals. During functional observational battery testing, significantly higher incidences of “possibly bloodstained” urine were observed for the high-dose animals at various times throughout the study; no other treatment-related changes were detected during the FOB assessment and there were no effects on motor activity. Reductions in mean body weights, body weight gains, and/or feed consumption values were seen in high-dose animals; there were no treatment-related effects on water consumption in any group. Moderate to large amounts of blood were detected in the urine of both sexes, primarily in the high-dose groups. Protein urinary concentrations were slightly increased in the high-dose animals and a dose-dependent decrease in urinary pH was observed in both sexes. No treatment-related changes in clinical chemistry, hematology, or cell morphology were observed. Organ weight changes were limited to statistically significantly lower mean absolute heart weights for high-dose males and lower mean terminal body weights and absolute thymus weights in high-dose females. Increases in a number of mean relative organ weights in high-dose females were considered related to the reduced terminal body weight changes and were not considered toxicologically significant. No toxicologically significant gross or microscopic lesions were observed at necropsy for any of the treated groups. Under the conditions of this study, the NOEL following exposure to 1,4-cyclohexanedimethanol in the drinking water for 13 weeks was 0.8% (resulting in doses of 479 mg/kg bw/day for male rats and 754 mg/kg bw/day for female rats).

Key value for chemical safety assessment

Toxic effect type:

concentration-driven

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Specific details on test material used for the study:

Test Substance Name: 1 ,4-Cyclohexanedimethanol (90%) in water ( 10%)
Synonym: CHDM-D90
HAELNo.: 99-0207
EANNo.: 907566
CAS No.: 000105-08-8
PM No.: 10674-00
SRIDNo.: 42490
Physical State and Appearance: Liquid, clear colorless
Source of Test Substance: Eastman Chemical Company, Kingsport, TN

Species:

rat

Strain:

Crj: CD(SD)

Remarks:

Sprague-Dawley rats [Crl:CD(SD)IGS BR]

Details on species / strain selection:

Rats were chosen for this study because they are a common representative species for oral and reproductive/developmental toxicity studies. They have a high fecundity, and this strain is routinely used in reproductive and developmental toxicity studies in our laboratory. Also, the rat is the primary rodent species recommended for use in the USEP A and OECD test guidelines.

Sex:

male/female

Details on test animals or test system and environmental conditions:

Test system

For both components of the study, male and nulliparous and non-pregnant female
Sprague-Dawley rats [Crl:CD(SD)IGS BR] were obtained from Charles River Laboratories, Stone Ridge (Kingston), NY. Twelve male rats per dose group were used for both portions of th study. These animals were 52 days of age and weighed 260.7 ± 12.5 grams (mean± SD) at the start of the study. Ten female rats per dose group were used exclusively for the 13-Week portion of the study. These animals were 53 days of age and weighed 187.8 ± 11.5 grams (mean± SD)
at the start of the study. Twelve female rats per dose group were used exclusively for the R/DTS portion of the study. These animals were 53 days of age and weighed 188.4 ± 13.7 grams (mean± SD) at the start of the study.

Rats were chosen for this study because they are a common representative species for oral and reproductive/developmental toxicity studies. They have a high fecundity, and this strain is routinely used in reproductive and developmental toxicity studies in our laboratory. Also, the rat is the primary rodent species recommended for use in the USEPA and OECD test guidelines.

Husbandry

Housing
Animals were housed in an Association for Assessment and Accreditation of Laboratory Animal Care International-accredited vivarium in accordance with the Guide for the Care and Use of Laboratory Animals (National Research Council, 1996). All animals were housed in suspended, stainless-steel mesh cages. Cages and racks were washed once a week. Absorbent paper, used to collect excreta, was changed daily. The female rats assigned exclusively to the 13-Week portion of the study (13-Week females) were singly housed for the duration of the study. The male rats and the female rats assigned exclusively to the RIDTS portion of the study (R/DTS females) were singly housed for the duration of the study except during the mating phase, during which time these animals were housed in pairs. No other studies were housed in the same rooms as this study.

Environmental Conditions

The study rooms were maintained at 21.4- 25.3°C and 30.4- 67.3% relative humidity for all but two days of the study. On Days 43 and 44, the Environmental Watchdog System was not activated for one of the rooms; therefore, there is no record of temperature or humidity for those days. However, the HVAC system in the vivarium displays an alarm whenever there is a deviation from the target values for temperature and humidity. There were no alarms to the HVAC on Days 43 and 44. Therefore, it is assumed that the temperature and humidity values were maintained at the target ranges of 22 ± 3°C and 30-70% relative humidity for these two days. A photoperiod of 12 hours light from 6am to 6 pm was maintained.

Route of administration:

oral: drinking water

Details on route of administration:

Test Substance Exposure
The rats were given drinking water containing 12.5, 8.0, 4.0 or 0.0 mg of the pure test substance per mL of drinking water (1.25, 0.80, 0.40 or 0.00%, respectively) ad libitum until they were euthanatized.

Preparation of Test Substance in Vehicle Mixtures
The test substance was received as a 90% solution in water. The amount of test substance used was adjusted for concentration prior to mixing with drinking water to yield concentrations of 12.5, 8.0 and 4.0 mg/mL of the active ingredient. Test substance in vehicle mixtures were prepared as needed, and used within 40 days based on the stability of the mixture.

Vehicle:

water

Details on oral exposure:

Stability of test substance in dosing solutions:
Stability in drinking water was determined by repeated analysis of aqueous solutions containing 1.0 and 12.5 mg/mL concentrations of the test substance. Mixtures were analyzed using gas chromatography with flame ionization detection (GC/FID). Refrigerated samples stored either in glass or Nalgene containers were analyzed on Days 0, 7, 14, 28 and 40. Room temperature samples stored in inverted glass containers sealed with a stopper and sipper tube were analyzed on Days 0, 1, 2, 5 and 7. Mixtures were stable under all tested conditions.

Exposure:
The male and female rats were exposed to CHDM in the drinking water ad libitum for 13 consecutive weeks until they were euthanized. The rats were offered drinking water containing 0, 4.0, 8.0 or 12.5 mg/mL of the test substance resulting in concentrations of 0, 0.4, 0.8, 1.25%, respectively. All surviving males were euthanized and necropsied on days 92 or 93; all surviving females were euthanized and necropsied on days 93 or 94.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

PURITY OF TEST SUBSTANCE:
The purity of the test substance was determined by GC/FID and Karl Fischer titration. Prior to use in the study, the test substance was determined to contain 90.0% by weight of 1,4-cyclohexanedimethanol, 9.79% by weight water, and 0.21% by weight of unknown materials. At the end of the study, the purity of the test article was determined to be 89.7% by weight of 1,4-cyclohexanedimethanol, 10.0% by weight water, and 0.30% by weight of unknown materials. Based on these data, the test substance was considered to be stable during the testing period.

STRUCTURE CONFIRMATION:
Structure was confirmed using gas chromatography with mass spectrometric detection (GS/MS). The mass spectrum of the test substance was consistent with the proposed structure based on comparison with the reference library spectrum of the test substance.

HOMOGENEITY OF DOSING SOLUTIONS:
Homogeneity of the test substance in drinking water was evaluated by measuring the concentration of the test substance at three levels (top, middle, and bottom of the mixing container) of all dosing mixtures using GC/FID.

The analytical concentrations of the test substance in the top, middle, and bottom layers for the 12.5 mg/mL mixture varied from 12.8 to 13.67 mg/mL ± 0.272. The analytical concentrations of the test substance in the top, middle, and bottom layers for the 8.0 mg/mL mixture varied from 7.191 to 7.735 mg/mL ± 0.144. The analytical concentrations of the test substance in the top, middle, and bottom layers for the 4.0 mg/mL mixture varied from 3.766 to 4.037 mg/mL ± 0.074. Based on the results, preparations were considered to be homogeneous.

CONCENTRATION OF TEST SUBSTANCE IN DOSING SOLUTIONS:
The concentration of the test substance in each batch of test mixture was determined prior to use by GC/FID analysis. The mean concentrations of the test substance varied between 97-102%, 93.4 - 108%, and 95.2-106% of the target concentrations of 4.0, 8.0, and 12.5 mg/mL, respectively.

Duration of treatment / exposure:

13 weeks

Frequency of treatment:

ad libitum in drinking water

Dose / conc.:

0.4 other: percent

Remarks:

Doses / Concentrations:
0.40% (4.0 mg/mL)
Basis:
nominal in water

Dose / conc.:

0.8 other: percent

Remarks:

Doses / Concentrations:
0.80% (8.0 mg/mL)
Basis:
nominal in water

Dose / conc.:

1.25 other: percent

Remarks:

Doses / Concentrations:
1.25 % (12.5 mg/mL)
Basis:
nominal in water

No. of animals per sex per dose:

12 males/group
10 females/group

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The exposure levels were based on two probe studies conducted in female rats. For both studies, the test substance was administered via drinking water for 7 consecutive days. Based on these study results, a concentration of 12.5 mg/mL (1.25%) was selected to maximize consumption of the test substance in drinking water. Concentrations of 8.0 mg/mL (0.80%) and 4.0 mg/mL (0.40%) were selected to provide evidence of possible dose-response relationships.

The animals were offered drinking water containing 1.25% (12.5 mg/mL), 0.80% (8.0 mg/ mL), 0.40% (4.0 mg/ mL) or 0.00% (0.0 mg/ mL) (Groups 4, 3, 2, and 1, respectively) of the active ingredient (1 ,4-Cyclohexanedimethanol) for approximately 80 - 96 days. Dose levels of the active ingredient achieved by this regimen were approximately 861,479, 256 and 0 mglkg body weight/day for the male rats, 1752, 754, 440 and 0 mg/kg body weight/day for the female rats.

During treatment, animals were monitored daily for changes in clinical signs, body weight, food and water consumption. At or around necropsy blood and urine were collected and animals were run through a functional observational battery. After necropsy, collected tissues were processed for histological examination.

Positive control:

no

Observations and examinations performed and frequency:

DAILY CLINICAL OBSERVATIONS:
Clinical examinations were conducted every morning for all animals except for days when functional observational battery occurred. Except for holidays, all animals were also observed for moribundity/mortality each weekday afternoon.

FUNCTIONAL OBSERVATIONAL BATTERY (FOB):
FOB conducted on all animals prior to study start and 1X/week through Week 13. Whenever possible, the FOB occurred at the same time each day. The observer was blind to treatment status and the same observer was used for each replicate throughout the study. Animals were observed for: severity and degree of lacrimation, salivation or discharges; piloerection and hair coat; pupillary size; exophthalmus; mucous membrane/skin color; unusual respiration; feces (amount and consistency); urine (amount and color); description of body position, coordination of movement, and gait abnormalities; description, incidence, and severity of convulsions and tremors; and stereotypy and bizarre behavior.

Additionally, prior to study start and during week 13 the animals were observed for: sensory function (vision and audition); proprioceptive reflex; and forelimb and hindlimb grip strength.

MOTOR ACTIVITY:
Motor activity was measured in 10-minute intervals for a total of 60 minutes during week 13 for all animals using an automated cage rack photobeam activity system (San Diego Instruments, San Diego, CA) attached to a Compaq 386SX computer. The total number of ambulations and total motor activity was calculated for the entire one-hour period.

BODY WEIGHTS:
Body weights were recorded for all animals on Days 0, 7, and at least weekly thereafter. The animals were fasted the day prior to necropsy and a terminal body weight was recorded at necropsy (measured after exsanguination but prior to necropsy).

FEED CONSUMPTION:
Feed consumption was measured on days 0, 7 and weekly thereafter except for weeks 9 and 10 when male feed consumption was not recorded due to the mating phase.

WATER CONSUMPTION:
Water consumption was measured on days 0, 4 and at least twice weekly thereafter except for weeks 9 and 10 when male water consumption was not recorded due to the mating phase.

OPTHALMIC EXAMINATION:
Prior to study start and on week 13, all rats were examined by a board certified ophthalmologist for retinal and corneal lesions using an indirect ophthalmoscope.

URINALYSIS:
On day 80 (male) and day 84 (female) rats were placed in metabolism cages for overnight urine collection. Urine was evaluated for color, clarity, output (mL/hr), specific gravity, pH, and the presence of glucose, protein and blood.

HEMATOLOGY AND CLINICAL CHEMISTRY:
All animals were fasted prior to necropsy, anesthetized with carbon dioxide, and blood was collected from the posterior vena cava. Blood was placed into vacutainer tubes and allowed to clot for serum analysis, tubes containing anticoagulant were used for analysis of whole blood samples, and blood smears were prepared for morphology and differential white blood cell counts.

Hematology tests included: hemoglobin concentration, hematocrit, red blood cell count, white blood cell count, red blood cell indices, prothrombin time, and platelet counts.

Clinical chemistry tests included: aspartate aminotransferase, alanine aminotransferase, sorbitol dehydrogenase, alkaline phosphatase, creatinine, urea nitrogen, glucose, total bilirubin, total protein, albumin, albumin/globulin ratio, total cholesterol, triglycerides, gamma-glutamyl transpeptidase, calcium, phosphorus, sodium, and potassium.

Sacrifice and pathology:

NECROPSY:
Following exsanguination, the rats were weighed and necropsied. The following tissues were fixed in 10% neutral buffered formalin: trachea, nasal passages, pharynx, larynx, lungs, aorta, heart, esophagus, stomach, duodenum, jejunem, ileum, cecum, colon, rectum, liver, pancreas, kidneys, urinary bladder, salivary glands, adrenal glands, thyroid gland, parathyroid glands, female mammary gland, pituitary gland, thymus, spleen, sternum (with bone marrow), mesenteric lymph nodes, cervical lymph nodes, brain (including sections of medulla/pons, cerebellar cortex, and cerebral cortex), sciatic nerve, spinal cord (including cervical, mid-thoracic and lumbar sections), skin, male accessory sex glands, ovaries, vagina, uterus, Fallopian tubes, and gross lesions. The eyes were fixed in Zenker’s solution for 4-6 hours and then rinsed overnight with slowly running water and stored in 70% ethyl alcohol. For the male rats, the right testis and right epididymis were fixed in Bouin’s solution after sperm motility analysis (details in a separate entry) and after approximately 24 hours, the tissues were rinsed twice with 50% ethyl alcohol and then stored in 70% ethyl alcohol. The left testis and left epididymis were placed in individual containers and frozen at -70°C.

ORGAN WEIGHTS:
The liver, kidneys, spleen, thymus, adrenal glands, heart, brain, testes, epididymides, ovaries, and uterus were weighed. Paired organs were weighed together.

HISTOPATHOLOGY:
All tissues (except for the left testis and left epididymis) from surviving male and female rats in the control and high-dose groups were examined for microscopic lesions. All tissues (except for the left testis and left epididimis) from male rats that died or were euthanized in extremis were examined microscopically. Additionally, all gross lesions from low- and mid-dose group animals were examined for microscopic lesions.

Statistics:

Mean values were calculated for body weight, feed consumption, total motor activity, total ambulations, grip strength, FOB behavior scores, serum chemistries, hematology values, organ weights, and organ-to-body weight ratios. Homogeneity of body weight, feed consumption, FOB behavior scores, motor activity, clinical pathology, and organ weight data were evaluated using Bartlett’s test (p≤0.01). Body weight, feed consumption, clinical pathology, and organ weight data were evaluated using a one-way analysis of variance (ANOVA) (p≤0.05) and Duncan’s multiple range test (p≤0.05). All other continuous data (grip strength, total motor activity, and total ambulations) and FOB behavior scores were evaluated using a repeated-measures/multivariate analysis of variance (p≤0.05). If significant time components were detected, then a one-way analysis of each test day was conducted. Post-hoc multiple comparison procedures such as Dunnett’s t-test were employed to identify the statistically significant means. Tests for linear trend over time in each group were conducted using linear regression (p≤0.05). If significant differences in total motor activity or total ambulations were seen, then the values for each ten minute interval were evaluated using a one-way analysis of variance (ANOVA) (p≤0.05) and Dunnett's t-test (p≤0.05). If the data were determined to be non-homogeneous, the data for that day were evaluated using the Kruskal-Wallis H-test (p≤0.05) followed by Mann-Whitney U-test (p≤0.05). Categorical FOB data were analyzed using a two-way or multiway frequency table analysis. If significant dose-behavior interactions were detected then a Fisher’s Exact test or Likelihood Ratio Chi-Square comparison was used to compare treated and control incidence at each weekly observation period.

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Observations related to test article exposure included hematuria and/or brown/red discoloration of urine first detected on day 6. All discolored urine tested positive for blood using an analytical reagent strip. This finding was observed exclusively in the high-dose animals (5 males, 4 females) except on days 8-9 when single male rats in the low- and mid-dose groups were observed with discolored urine. Softened or reduced feces were observed in a few animals in all groups, including controls. A single incidence of minimal dehydration was observed for one male and female in the high-dose group.

Additional observations present in a few high-dose animals included hypothermia, pallor, partially opened eyes, piloerection, unkempt haircoat and brown discoloration of the skin of the tail and inguinal or abdominal hair. No other test substance-related clinical abnormalities were observed.

Mortality:

mortality observed, treatment-related

Description (incidence):

One male rat from the low-dose group was found dead on day 12 and another male rat in the high-dose group was euthanized in extremis during the study on day 39.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

In males, mean body weights were significantly lower (p≤0.05) for the high-dose group during Weeks 1-7 and Week 9 when compared to control. Mean body weight gains were significantly less during Weeks 1, 2, and 5 for the high-dose animals, and higher during Week 7 for the mid-dose animals and Week 10 for the high-dose animals. There were no differences in overall study mean body weight change for any dose group compared to the control.

High-dose females had significantly lower (p≤0.05) mean body weights from Week 3 through termination and mean body weight changes were also significantly lower as compared to control during Week 1.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Mean feed consumption was significantly lower (p≤0.05) for high-dose males during Weeks 1, 2, 4, and 5, and for high-dose females during Weeks 1, 3-9, and 11-13 when compared to control.

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

no effects observed

Description (incidence and severity):

There were no effects in male rats. Mean values were significantly lower (p≤0.05) for mid-dose females on Days 42, 67, 74, and 77 when compared to control but this change was not considered to be treatment-related since it was not observed in animals from the high-dose group.

Ophthalmological findings:

no effects observed

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

Effects in males consisted of increased percentages of metamyelocytes in the high-dose group and decreased white blood cells counts in low-dose animals. Changes in females consisted of increased red blood cells counts and hematocrit values for mid-dose animals and decreased red blood cell counts for the high-dose animals. No overt changes in hematology values were observed for the one male rat in the high-dose group that was euthanized in extremis. Minimal to minor abnormalities which occurred with greater frequency in treated groups included macrocytosis, anisocytosis, and polychromia.

Minimal to minor changes in cell morphology including poikilocytosis, microcytosis, spherocytosis, hypochromasia, target cells, and Howell-Jolly bodies occurred sporadically (1-2 animals/group) or occurred with approximately equal frequencies between the control and treated groups, indicating that only a few cells were atypical.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Changes in clinical chemistry parameters included increased (p≤0.05) concentrations of urea nitrogen, creatinine, and cholesterol for low-dose males and increased cholesterol concentrations for high-dose females. For the one male rat in the high-dose group that was euthanized in extremis, clinical chemistry changes included elevated levels of creatinine, gamma-glutamyl transpeptidase, and cholesterol.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

Descriptions provided in the RSS for the 90-day study as this was a 90-day repeart dose with the reproductive screenTraces of blood were detected in the urine from 2-3 male rats per group (common for males rats of this strain and age) while moderate to large amounts of blood were detected in 3 male rats from the high-dose group and one male in the low-dose group. Large amounts of blood were also detected in the urine from 3 females from the high-dose group. A slight dose-dependent increase in urinary protein concentration was observed with control and low-dose values ranging from a trace to 30 mg/dl, mid-dose group values generally between 30-100 mg/dl, and 62% of the high-dose animals with values between 100 mg/dl and 300 mg/dl (10 rats) or > 2000 mg/dl (3 rats). A dose-dependent decrease in urinary pH was also observed with values between 7.0-8.0 for control, 6.0-7.0 for low-dose animals, 6.0-6.5 for mid-dose animals, and 6.0 for high-dose animals. There were no adverse effects on glucose or specific gravity values, volume or clarity.

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

Functional Observational Battery:
Significantly greater (p≤0.05) incidences of “possibly bloodstained” urine were observed for the high-dose males during Weeks 2, 3, 4, and 7 of the study. This finding was also observed in high-dose females during Weeks 6-11; however these occurrences did not become statistically significant (p≤0.05) until Weeks 12 and 13.

Motor Activity:
There were no significant differences in mean total ambulations or mean total motor activity counts for any groups.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Organ weight changes were limited to lower (p≤0.05) mean absolute heart weights for high-dose males when compared to control. Lower mean terminal body weights and absolute thymus weights, and higher mean relative liver, kidney, heart, brain, and adrenal gland weights were observed in the high-dose females when compared to control.

Gross pathological findings:

no effects observed

Description (incidence and severity):

Descriptions provided in the RSS for the 90-day study as this was a 90-day repeart dose with the reproductive screen

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

There were several microscopic lesions observed in all groups, including controls, but these lesions were considered to be incidental in nature and not related to test article administration.

Histopathological findings: neoplastic:

no effects observed

Other effects:

not specified

Details on results:

Doses received:
Mean daily doses of the test substance received for the 0.0, 4.0, 8.0, and 12.5 mg/mL groups were approximately 0, 256, 479, and 861 mg/kg bw/day for males and 0, 440, 754, and 1752 mg/kg bw/day for females.

Mortality:
One male rat from the low-dose group was found dead on day 12 and another male rat in the high-dose group was euthanized in extremis during the study on day 39.

Clinical Observations:
Observations related to test article exposure included hematuria and/or brown/red discoloration of urine first detected on day 6. All discolored urine tested positive for blood using an analytical reagent strip. This finding was observed exclusively in the high-dose animals (5 males, 4 females) except on days 8-9 when single male rats in the low- and mid-dose groups were observed with discolored urine. Softened or reduced feces were observed in a few animals in all groups, including controls. A single incidence of minimal dehydration was observed for one male and female in the high-dose group.

Additional observations present in a few high-dose animals included hypothermia, pallor, partially opened eyes, piloerection, unkempt haircoat and brown discoloration of the skin of the tail and inguinal or abdominal hair. No other test substance-related clinical abnormalities were observed.

Functional Observational Battery:
Significantly greater (p≤0.05) incidences of “possibly bloodstained” urine were observed for the high-dose males during Weeks 2, 3, 4, and 7 of the study. This finding was also observed in high-dose females during Weeks 6-11; however these occurrences did not become statistically significant (p≤0.05) until Weeks 12 and 13.

Motor Activity:
There were no significant differences in mean total ambulations or mean total motor activity counts for any groups.

Body Weights:
In males, mean body weights were significantly lower (p≤0.05) for the high-dose group during Weeks 1-7 and Week 9 when compared to control. Mean body weight gains were significantly less during Weeks 1, 2, and 5 for the high-dose animals, and higher during Week 7 for the mid-dose animals and Week 10 for the high-dose animals. There were no differences in overall study mean body weight change for any dose group compared to the control.

High-dose females had significantly lower (p≤0.05) mean body weights from Week 3 through termination and mean body weight changes were also significantly lower as compared to control during Week 1.

Feed Consumption:
Mean feed consumption was significantly lower (p≤0.05) for high-dose males during Weeks 1, 2, 4, and 5, and for high-dose females during Weeks 1, 3-9, and 11-13 when compared to control.

Water Consumption:
There were no effects in male rats. Mean values were significantly lower (p≤0.05) for mid-dose females on Days 42, 67, 74, and 77 when compared to control but this change was not considered to be treatment-related since it was not observed in animals from the high-dose group.

Ophthalmic Examination:
No treatment-related ophthalmologic changes were detected.

Urinalysis:
Traces of blood were detected in the urine from 2-3 male rats per group (common for males rats of this strain and age) while moderate to large amounts of blood were detected in 3 male rats from the high-dose group and one male in the low-dose group. Large amounts of blood were also detected in the urine from 3 females from the high-dose group. A slight dose-dependent increase in urinary protein concentration was observed with control and low-dose values ranging from a trace to 30 mg/dl, mid-dose group values generally between 30-100 mg/dl, and 62% of the high-dose animals with values between 100 mg/dl and 300 mg/dl (10 rats) or > 2000 mg/dl (3 rats). A dose-dependent decrease in urinary pH was also observed with values between 7.0-8.0 for control, 6.0-7.0 for low-dose animals, 6.0-6.5 for mid-dose animals, and 6.0 for high-dose animals. There were no adverse effects on glucose or specific gravity values, volume or clarity.

Hematology:
Effects in males consisted of increased percentages of metamyelocytes in the high-dose group and decreased white blood cells counts in low-dose animals. Changes in females consisted of increased red blood cells counts and hematocrit values for mid-dose animals and decreased red blood cell counts for the high-dose animals. No overt changes in hematology values were observed for the one male rat in the high-dose group that was euthanized in extremis. Minimal to minor abnormalities which occurred with greater frequency in treated groups included macrocytosis, anisocytosis, and polychromia.

Minimal to minor changes in cell morphology including poikilocytosis, microcytosis, spherocytosis, hypochromasia, target cells, and Howell-Jolly bodies occurred sporadically (1-2 animals/group) or occurred with approximately equal frequencies between the control and treated groups, indicating that only a few cells were atypical.

Clinical chemistry:
Changes in clinical chemistry parameters included increased (p≤0.05) concentrations of urea nitrogen, creatinine, and cholesterol for low-dose males and increased cholesterol concentrations for high-dose females. For the one male rat in the high-dose group that was euthanized in extremis, clinical chemistry changes included elevated levels of creatinine, gamma-glutamyl transpeptidase, and cholesterol.

Organ weights:
Organ weight changes were limited to lower (p≤0.05) mean absolute heart weights for high-dose males when compared to control. Lower mean terminal body weights and absolute thymus weights, and higher mean relative liver, kidney, heart, brain, and adrenal gland weights were observed in the high-dose females when compared to control.

Gross Pathology:
Gross lesions were observed in all groups, including controls, but none of the gross lesions were considered related to test article administration.

Histopathology:
There were several microscopic lesions observed in all groups, including controls, but these lesions were considered to be incidental in nature and not related to test article administration.

Dose descriptor:

NOEL

Effect level:

0.8 other: %

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Critical effects observed:

not specified

Stability of test substance:

For refrigerated samples stored in glass containers, concentrations of test substance in the 12.5 and 1.0 mg/mL solutions were 12.5 ± 0.25 and 0.949 ± 0.0194 prior to storage and 11.1 ± 0.26 and 0.873 ± 0.0196 after 40 days of storage. For refrigerated samples stored in Nalgene containers, concentrations of test substance were 11.6 ± 0.13 and 0.960 ± 0.0146 prior to storage and 11.2 ± 0.08 and 0.910 ± 0.0264 after 40 days of storage. For samples stored in glass containers at room temperature, concentrations of test substance were 12.2 ± 0.24 and 0.980 ± 0.0183 prior to storage and 12.5 ± 0.14 and 0.987 ± 0.0133 after 7 days of storage. Based on these results, the refrigerated mixtures were stored in either Nalgene or glass containers during the study, and the room temperature mixtures were stored in glass containers during the study.

Probe Studies:

In the first study, 12 female rats were offered drinking water containing 0 mg/mL (0%), 5.0 mg/mL (0.50%), 10.0 mg/mL (1.00%), or 15.0 mg/mL (1.50%) of 1,4-cyclohexanedimethanol for seven days. No mortality was observed in the study; animals appeared clinically normal; slight decreases in body weight were reported for the first 3 days of the study but all animals had gained weight by Day 7; and there were no effects on feed consumption. Water consumption in the high-dose group was 50% of the control, possibly because the animals found the water unpalatable. Therefore, a second probe study was conducted to determine the maximum dose possible by the drinking water route.

For the second study, six females were offered drinking water containing either 0.0 or 12.5 mg/mL (0 and 1.25%, respectively) of the test substance for seven days. No mortality was observed; clinical observations were normal; slight decreases in body weight were reported for the first 4 days of the study but all animals had gained weight by Day 7. Significant decreases in food consumption were noted on Day 2 for the treated group; and water consumption was comparable among the treated and control groups. The animals treated with concentrations of 12.5 mg/mL received the largest dose in the probe study so this concentration was chosen to achieve a target dose level of 1000 mg/kg bw/day for the 13-week study.

Conclusions:

In a subchronic oral toxicity study, 1,4-cyclohexanedimethanol was administered to 10 Crl:CD(SD)IGS BR females/dose and 12 Crl:CD(SD)IGS BR males/dose via the drinking water at dose concentrations of 0 (0 mg/mL), 0.4% (4.0 mg/mL), 0.8% (8.0 mg/mL), and 1.25% (12.5 mg/mL) for 13 weeks. The NOEL for systemic toxicity was determined to be 0.8% (8.0 mg/mL) based upon mortality, abnormalities in the urine and feces, reductions in body weight and body weight gain, decreased feed consumption, and increased urinary protein in animals exposed to 1.25 % (12.5 mg/mL).

No target organ effects were observed in male or female rats following ad libitum exposure to up to 0.8% 1,4-cyclohexanedimethanol in the drinking water (resulting in doses of 479 mg/kg bw/day for males and 754 mg/kg bw/day for females) for 13 weeks. The primary adverse effect observed in the drinking water study was minimal to severe hematuria, primarily in animals receiving 1.25% test material in the drinking water, but there were no corresponding adverse effects on hematology or gross or microscopic effects on any target organ including kidneys and urinary bladder. The NOEL for systemic effects in both sexes in the drinking water study was considered to be 0.8% (resulting in doses of 479 and 754 mg/kg bw/day in male and female rats, respectively). Based on the results of this study, 1,4-cyclohexanedimethanol is not classified for “Specific Target Organ Toxicity – Repeated Exposure” according to GHS.

Executive summary:

In a subchronic oral toxicity study, 1,4-cyclohexanedimethanol was administered via the drinking water to groups of male and female Crl:CD(SD)IGS BR rats (10 females/dose and 12 males/dose) at target concentrations of 0 (0 mg/mL), 0.4% (4.0 mg/mL), 0.8% (8.0 mg/mL), and 1.25% (12.5 mg/mL) for 13 weeks. One male rat each from the low- and high-dose groups were found dead or euthanized in extremis before study termination. Clinical abnormalities related to test substance exposure included minimal to severe hematuria and/or brown/red discoloration of the urine, primarily in high-dose group animals. Softened or reduced feces occurred in animals in all groups but severity and incidence were slightly greater among treated groups. Other abnormal signs were observed occasionally in a few treated animals. During functional observational battery testing, significantly higher incidences of “possibly bloodstained” urine were observed for the high-dose animals at various times throughout the study; no other treatment-related changes were detected during the FOB assessment and there were no effects on motor activity. Reductions in mean body weights, body weight gains, and/or feed consumption values were seen in high-dose animals; there were no treatment-related effects on water consumption in any group. Moderate to large amounts of blood were detected in the urine of both sexes, primarily in the high-dose groups. Protein urinary concentrations were slightly increased in the high-dose animals and a dose-dependent decrease in urinary pH was observed in both sexes. No treatment-related changes in clinical chemistry, hematology, or cell morphology were observed. Organ weight changes were limited to statistically significantly lower mean absolute heart weights for high-dose males and lower mean terminal body weights and absolute thymus weights in high-dose females. Increases in a number of mean relative organ weights in high-dose females were considered related to the reduced terminal body weight changes and were not considered toxicologically significant. No toxicologically significant gross or microscopic lesions were observed at necropsy for any of the treated groups.

Under the conditions of this study, the NOEL following exposure to 1,4-cyclohexanedimethanol in the drinking water for 13 weeks was 0.8% (resulting in doses of 479 mg/kg bw/day for male rats and 754 mg/kg bw/day for female rats).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

754 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

The NOAEL findings were 754 mg/kg/day for females and 479 mg/kg/day for males. The time per week box can not be populated as the study was in drinking water.

System:

other: none

Organ:

not specified

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Mode of Action Analysis / Human Relevance Framework

none

Additional information

The potential for 1,4-cyclohexanedimethanol to cause target organ toxicity following repeated exposure is well understood. One guideline repeat-exposure drinking water study and a lesser non-guideline feeding study were available for review. In a subchronic oral toxicity study with separate reproduction/developmental toxicity screening test conducted according to OECD Guideline 408, male and female Sprague Dawley rats were exposed to up to 12.5 mg/mL (1.25%) of 1,4-cyclohexanedimethanol ad libitum in the drinking water for 13 weeks. Test substance-related clinical abnormalities included minimal to severe hematuria and/or brown/red discoloration of the urine, primarily in high-dose group animals of both sexes. During functional observational battery testing, significantly higher incidences of “possibly bloodstained” urine were reported for high-dose group animals but no other treatment-related changes were detected during the FOB assessment and there were no effects on motor activity. Reductions in mean body weights, body weight gains, and/or feed consumption values were seen in high-dose group animals but there were no effects on water consumption in any group. There were no treatment-related effects on clinical chemistry, hematology, or cell morphology. Although statistically significant lower mean absolute heart weights were seen in high-dose group males and lower mean terminal body weights and absolute thymus weights were seen in high-dose group females, no toxicologically significant gross or microscopic lesions were observed at necropsy for any of the treated groups. Under the conditions of this study, the NOEL following exposure to 1,4-cyclohexanedimethanol in the drinking water for 13 weeks was 0.8% (resulting in doses of 479 mg/kg bw/day for male rats and 754 mg/kg bw/day for female rats.)

In a supporting subchronic toxicity study in which groups of 5 rats/sex were administered up to 1.0% 1,4-cyclohexanedimethanol ad libitum in the basal diet for 36 days, there were no dose-related statistically significant effects on mortality, body weight, food consumption and feed efficiency, hematology, clinical chemistry, urinalysis, organ weights or gross or microscopic pathology in either sex. Under the conditions of this study, the NOAEL following exposure to 1,4-cyclohexanedimethanol in the feed for 36 days was 1.0% in male and female rats.

Justification for classification or non-classification

No target organ effects were observed in male or female rats following ad libitum exposure to up to 0.8% 1,4-cyclohexanedimethanol in the drinking water (resulting in doses of 479 mg/kg bw/day for males and 754 mg/kg bw/day for females) for 13 weeks or at up to 1% in the diet for 36 days. The primary adverse effect observed in the drinking water study was minimal to severe hematuria, primarily in animals receiving 1.25% test material in the drinking water, but there were no corresponding adverse effects on hematology or gross or microscopic effects on any target organ including kidneys and urinary bladder. The NOEL for systemic effects in both sexes in the drinking water study was considered to be 0.8% (resulting in doses of 479 and 754 mg/kg bw/day in male and female rats, respectively). No adverse effects were observed on mortality, body weight, food consumption and feed efficiency, hematology, clinical chemistry, urinalysis, organ weights or gross or microscopic pathology in either sex in the 36-day feeding study and the NOAEL was considered to be 1.0%. Based on a weight-of-the-evidence assessment, 1,4-cyclohexanedimethanol is not classified for “Specific Target Organ Toxicity – Repeated Exposure” according to GHS.